Activation of adenosine A2A receptors induces SUMOylation of HIF-1α.

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Hypoxia is an established driver of numerous deleterious cell processes, including immunosuppression and tumor growth. Hypoxia-inducible factor 1α (HIF-1α) accumulates during periods of sustained hypoxia due to the action of multiple post-translational modifications, including the Small Ubiquitin-like modifier (SUMO) pathway. Although enhanced activity of the SUMO-pathway is upregulated during hypoxia, the mechanistic basis for this effect remains unclear. It has been proposed that the Gαs protein-coupled adenosine A2A receptor (A2AR), which is also activated during sustained hypoxia, mediates SUMOylation at a number of target proteins. We sought to determine if activation of A2AR by hypoxia induces SUMOylation of HIF-1α and which of three potential SUMO-sites on HIF-1α are modified by this mechanism. HEK293T cells expressing A2AR and treated with an A2AR agonist (CGS) showed significantly increased HIF-1α accumulation compared to untreated controls after exposure to hypoxia. Similarly, CGS and hypoxia enhanced SUMOylation of HIF-1α in human Jurkat T cells. Using site-directed mutagenesis and Förster resonance energy transfer (FRET), we determined that A2AR-mediated SUMOylation of HIF-1α occurs at lysine 532. Finally, we show that (1) this mechanism of regulation increases the mRNA levels of multiple HIF-1α transcriptional targets; and (2) this mechanism can be precluded by treating cells with a small molecule inhibitor of the SUMO pathway. Together, these data highlight a central role for A2AR-mediated SUMOylation of HIF-1α in mediating the effects of chronic hypoxia in Jurkat T cells.